TR-USAXS/SAXS investigation of Myelin Basic Protein liquid-liquid phase separation kinetics
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Liquid-liquid phase separation (LLPS) of intrinsically disordered proteins has attracted a wide attention in the biological and biophysical community. We suggest to investigate the kinetics of the LLPS of the intrinsically disordered myelin basic protein (MBP) and connected structural changes of the protein using time-resolved (TR) USAXS/ SAXS at ID02 with the stopped flow technique. MBP is essential for the integrity of myelin sheaths ensuring flawless neuronal signal propagation and it is assumed that LLPS is necessary for its physiological function. We propose to use 3 detector distances to investigate a q-range of 1∙10^-3 - 5∙10^0 1/nm in order to observe internal protein structure, droplet nucleation and droplet growth evolution. Our proposed experiments take 60 hours of pure X-ray beamtime, so we believe that 3 days of beamtime are necessary considering extra time required to set up the device, do cleaning, test-shots, find suitable exposure times and radiation intensities.
内在无序蛋白（intrinsically disordered proteins）的液-液相分离（Liquid-liquid phase separation, LLPS）已在生物与生物物理学界受到广泛关注。本研究拟针对内在无序髓鞘碱性蛋白（myelin basic protein, MBP）的液-液相分离动力学及其伴随的蛋白质结构变化展开研究，将在ID02线站借助停流技术，采用时间分辨超小角X射线散射/小角X射线散射（time-resolved, TR USAXS/SAXS）开展实验。MBP对维持髓鞘的完整性、保障神经元信号的正常传导至关重要，且学界普遍认为液-液相分离对其生理功能具有必要性。本实验拟设置3个探测距离，以覆盖1×10^-3 至 5×10^0 每纳米的q范围，从而实现对蛋白质内部结构、液滴成核及液滴生长演化过程的观测。本次实验所需纯X射线束时为60小时，考虑到装置搭建、设备清洁、测试曝光、确定合适的曝光时长与辐射强度等额外耗时，我们认为需申请3天的束时资源。
